Pilot Phase I/II Personalized Therapy Trial for Metastatic Colorectal Cancer: Evaluating the Feasibility of Protein Pathway Activation Mapping for Stratifying Patients to Therapy with Imatinib and Panitumumab

Abstract

This nonrandomized phase I/II trial assessed the efficacy/tolerability of imatinib plus panitumumab in patients affected by metastatic colorectal cancer (mCRC) after stratification to treatment by selection of activated imatinib drug targets using reverse-phase protein array (RPPA). mCRC patients presenting with a biopsiable liver metastasis were enrolled. Allocation to the experimental and control arms was established using functional pathway activation mapping of c-Kit, PDGFR, and c-Abl phosphorylation by RPPA. The experimental arm received run-in escalation therapy with imatinib followed by panitumumab. The control arm received panitumumab alone. Seven patients were enrolled in the study. For three of the seven patients, sequential pre- and post-treatment biopsies were used to evaluate the effect of the therapeutic compounds on the drug targets and substrates. A decrease in the activation level of the drug targets and downstream substrates was observed in two of three patients. Combination therapy increased the activation of the AKT-mTOR pathway and several receptor tyrosine kinases. This study proposes a novel methodology for stratifying patients to personalized treatment based on the activation level of the drug targets. This workflow provides the ability to monitor changes in the signaling pathways after the administration of targeted therapies and to identify compensatory mechanisms.

Phosphotyrosine-based-phosphoproteomics scaled-down to biopsy level for analysis of individual tumor biology and treatment selection

“…On this latter point, kinase-substrate analysis (KSEA) [34] and integrated personalized signatures (pCHIPS) [35] have recently been proposed for kinase pathway activity or drug prioritization, but these methods have not been validated for single clinical samples. Antibody-based protein arrays may provide a more high-throughput alternative phosphoproteomics approach and have shown potential for the identification of treatment targets [36] and patient stratification [37]. Drawbacks of this approach are the limited number of phosphosite-specific antibodies targeting assumed epitopes of interest [20,38].…”

Reverse Phase Protein Arrays: Mapping the Path Towards Personalized Medicine

“…Jameson et al recently reported a turn-around-time of 12–23 days (median 15.5 days) in a multi-site, multi-omic clinical trial (Side-Out, Table 1) in which fresh biopsies were subjected to immunohistochemistry, cDNA microarray analysis, and LCM prior to RPPA [5]. The turnaround-time for LCM-RPPA of clinical biopsy samples in our clinical trials ranges from 3 – 10 business days from biopsy to report [5, 22] [Dr. M. Pierobon, 8 Oct 2014, written personal communication]. Microdissection of heterogeneous tissue samples is required to obtain meaningful information that can reveal unique protein pathway signatures of a patient’s tumor, which in turn can be exploited as therapeutic targets.…”

“…Nonetheless, new molecular targets and combination therapies are actively being evaluated in clinical trials, expediting the translation of basic research into clinical applications (Table 1). RPPA analysis includes the following functional information about the state of actionable drug targets: a) protein signal pathway network analysis, b) upstream/downstream linkage analysis, c) protein signaling across classes of samples/treatments, d) predictive treatment efficacy and patient stratification, and e) post-translational proteomic data [5, 20–22]. This data is unattainable by genomic and transcriptomic analyses.…”

“…Validation entails assessing RPPA performance following strict standard operating procedures with on-going documentation of performance parameters such as inter-assay precision, quality control ranges, and adequacy and performance of calibrators over time [5, 22]. High throughput quantitative proteomic techniques, such as reverse-phase protein arrays in which 80 or more samples can be assessed on the same array under the same experimental conditions, have made the analysis of more complex biological systems achievable.…”

“…The RPPA platform allows the profiling and comparison of the functional state of protein signaling pathways, over time, to follow the course of disease as well as its response to therapy. Characterization of novel drug targets, and the discovery of potential diagnostic and prognostic biomarkers are placing reverse phase protein array technology as the preferred platform for proteomic cancer studies [1, 2, 10, 22–27]. However, RPPA have been utilized to assess the state of signaling kinases in non-neoplastic conditions, such as during Rift Valley Fever (RFV) infection and in ophthalmologic conditions such as macular degeneration [28, 29].…”